Remodeling of regulation of cell growth by glycogene and its application.

糖原对细胞生长调控的重塑及其应用。

• 批准号: 09680590
• 负责人: FUJII Junichi
• 金额: $ 2.18万
• 依托单位: Osaka University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1997
• 资助国家: 日本
• 起止时间: 1997 至 1998
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09680590/
• 关键词: Apoptosis; Growth factor; Signal transduction; EGF receptor; EGF-受容体

Although the importance of glycosylation of components on the cell membrane is widely accepted, the relationship with cell growth has been only poorly investigated.We remodeled the sugar chains on the cells by transfecting PC 12 cells with rat beta- 1, 4-N- acetlylglucosamyltransferase III gene (GnT III) and effects of them on the cellular responses to the nerve growth factor (NGF) were examined.The results show that branch structure, a product of the gene, was found in GnT III-transfected PC 12 cells.However, no change in the growth rate or neurites formation was observed.When effects of NGF on Trk, a NGF receptor, was investigated, tyrosine phosphotylation, which occurs in control cells, was not observed.Moreover, dimerization essential for signal transduction to nucleus did not occur.Then we transfected B 16 melanoma, a line commonly used for metastasis, NIH3T3, and CHOK1 cells with EGF receptor and GnT III cDNAs by electroporation and selected neomycin-resistant cells.Several clones which highly express GnT III were isolated.The sugar chains of the EGF receptor was modified with bisecting GlcNAc structure and dysfunction of cellular response to EGF stimuli was observed.Currently we are investigating the MAP kinase cascade downstream of the EGF receptor signaling.

尽管细胞膜成分糖基化的重要性已被广泛接受，但对其与细胞生长的关系的研究还很少。我们用大鼠β-1，4-N-乙酰氨基葡萄糖转移酶III基因(GnT III)来重塑PC 12细胞的糖链，并观察它们对细胞对神经生长因子(NGF)反应的影响。结果表明，在GnT III转基因的PC 12细胞中发现了该基因的产物--分支结构。然而，NGF对神经生长因子受体Trk的影响并未观察到生长速度或神经突的形成。当NGF对神经生长因子受体Trk的影响时，酪氨酸磷酸化，然后，我们通过电穿孔的方法将EGF受体和GnT III cDNAs导入B16黑色素瘤细胞、NIH3T3细胞和CHOK1细胞，并筛选出了几个高表达GnT III的细胞克隆。用一分为二的GlcNAc结构修饰了EGF受体的糖链，并观察到细胞对EGF刺激的反应障碍。目前，我们正在研究EGF受体下游的MAP激酶级联反应。

项目成果

Y.Miyazaki: "Gastrin induces heparin-binding epidermal growth factor-like growth factor in rat gastric epithelial cells transfected with gastrin receptor." Gastroenterology. 116. 78-89 (1999)

Y.Miyazaki：“胃泌素在转染胃泌素受体的大鼠胃上皮细胞中诱导肝素结合表皮生长因子样生长因子。”

N.Miyazawa: "Immunological detection of fructated proteins in in vitro and in vivo." Biochem J. 336. 101-107 (1998)

N.Miyazawa：“体外和体内果糖蛋白的免疫学检测。”

Y.Kawasaki: "Specific detection of the early process of the glycation reaction by fructose and glucose in diabetic rat lens." FEBS Lett. 441. 116-120 (1998)

Y.Kawasaki：“糖尿病大鼠晶状体中果糖和葡萄糖糖化反应早期过程的特异性检测。”

M.Asahi: "The Oxidation of Selenocysteine is Involved in the Inactivation of Glutathione Peroxidase by Nitric Oxide Donor" J.Biol.Chem.272. 19152-19157 (1997)

M.Asahi：“硒代半胱氨酸的氧化参与一氧化氮供体对谷胱甘肽过氧化物酶的失活”J.Biol.Chem.272。

J.Fujii: "A defect in the mitochondrial import of mutant Mn-superoxide dismutase produced in Sf21 cells." J Biochem. 124. 340-346 (1998)

J.Fujii：“Sf21 细胞中产生的突变锰超氧化物歧化酶的线粒体输入存在缺陷。”